Parichy David M, Turner Jessica M, Parker Nathan B
Section of Integrative Biology, Section of Molecular, Cell and Developmental Biology, Institute for Cellular and Molecular Biology, University of Texas at Austin, 1 University Station, C0930, Austin, TX 78712, USA.
Dev Biol. 2003 Apr 15;256(2):221-41. doi: 10.1016/s0012-1606(03)00016-2.
Multipotent neural crest stem cells have been identified in late gestation amniote embryos. Yet, significant questions remain about the mechanisms by which these cells are generated, maintained, and recruited during postembryonic development. The zebrafish, Danio rerio, offers an opportunity to identify genes essential for these processes, by screening for mutants with defects in traits likely to depend on these cells during metamorphosis and adult life. One such trait is the pigment pattern formed by neural crest-derived pigment cells, or chromatophores, which include black melanophores, yellow xanthophores, and iridescent iridophores. Previous analyses have demonstrated that the adult zebrafish pigment pattern depends on the de novo differentiation of latent precursor cells during both early and late phases of pigment pattern metamorphosis. To better understand the development of these cells, in this study, we analyze the zebrafish puma mutant, which ablates most of the adult melanophores that differentiate during metamorphosis, but leaves intact early larval melanophores that differentiate during embryogenesis. We use epistasis analyses to show that puma promotes the development of both early-appearing metamorphic melanophores that depend on the kit receptor tyrosine kinase, as well as late-appearing metamorphic melanophores that depend on both the G-protein-coupled endothelin receptor b1 (ednrb1) and the kit-related fms receptor tyrosine kinase. We further demonstrate that, during pigment pattern metamorphosis, puma mutants have deficiencies in the numbers of cells expressing transcripts for kit, ednrb1, and fms, as well as the HMG domain transcription factor sox10. Because the puma mutant phenotype is temperature-sensitive, we use temperature-shift experiments to identify a critical period for puma activity during pigment pattern metamorphosis. Finally, we use cell transplantations to show that puma acts cell-autonomously to promote the expansion of pigment cell lineages during metamorphosis. These results suggest a model for the lineage diversification of neural crest stem cells during zebrafish postembryonic development.
多能神经嵴干细胞已在妊娠晚期的羊膜动物胚胎中被鉴定出来。然而，关于这些细胞在胚胎后发育过程中产生、维持和募集的机制，仍存在重大问题。斑马鱼（Danio rerio）为鉴定这些过程所必需的基因提供了一个机会，通过筛选在变态发育和成年期可能依赖这些细胞的性状存在缺陷的突变体。其中一个这样的性状是由神经嵴衍生的色素细胞或色素体形成的色素模式，色素体包括黑色的黑素细胞、黄色的黄色素细胞和彩虹色的虹彩细胞。先前的分析表明，成年斑马鱼的色素模式依赖于色素模式变态发育早期和晚期潜在前体细胞的从头分化。为了更好地理解这些细胞的发育，在本研究中，我们分析了斑马鱼puma突变体，该突变体消除了变态发育期间分化的大多数成年黑素细胞，但保留了胚胎发育期间分化的早期幼虫黑素细胞。我们使用上位性分析表明，puma促进了依赖于kit受体酪氨酸激酶的早期出现的变态黑素细胞以及依赖于G蛋白偶联内皮素受体b1（ednrb1）和kit相关fms受体酪氨酸激酶的晚期出现的变态黑素细胞的发育。我们进一步证明，在色素模式变态发育期间，puma突变体在表达kit、ednrb1和fms转录本以及HMG结构域转录因子sox10的细胞数量上存在缺陷。由于puma突变体表型是温度敏感的，我们使用温度转换实验来确定色素模式变态发育期间puma活性的关键时期。最后，我们使用细胞移植表明，puma在变态发育期间自主发挥作用，促进色素细胞谱系的扩展。这些结果提示了斑马鱼胚胎后发育过程中神经嵴干细胞谱系多样化的模型。
